The effects of U46619, a thromboxane mimic, on cytosolic Ca++ concentration and platelet aggregation were determined in human platelets. Cytosolic Ca++ concentration was determined via Quin-2 fluorescence and platelet aggregation studied in an aggregometer. Addition of U46619 to the platelet suspension produced a sharp increase in cytosolic Ca++ and platelet aggregation. Pretreatment of platelets with prostaglandin I2, PGD2, PGE1, PGF2alpha, dibutyryl2 cyclic AMP or forskolin prevented the increase in cytosolic Ca++ and the associated platelet aggregation induced by U46619. In platelets treated with PGE2, 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate hydrochloride (TMB-8), or verapamil (V), U46619 produced a much slower increase in cytosolic Ca++. Although cytosolic Ca++ concentration eventually reached values equal to, or greater than, those of controls, no platelets aggregation was observed. These data suggest that U46619 induces platelet aggregation through an increase in cytosolic CA++, and that both CA++ entry and its release from intracellular storage sites probably contribute to the increase in cytosolic Ca++. Furthermore, the rate of the increase and the magnitude of the rise in cytosolic Ca++ concentration appear to be crucial in platelet aggregation induced by U46619. Our data also suggest that PGs inhibit U46619-induced platelet aggregation by preventing the increase in cytosolic Ca++, and these effects are probably mediated via an increase in cAMP.